Melting of a β-Hairpin Peptide Using Isotope-Edited 2D IR Spectroscopy and Simulations

Abstract

Isotope-edited two-dimensional infrared spectroscopy has been used to characterize the conformational heterogeneity of the beta-hairpin peptide TrpZip2 (TZ2) across its thermal unfolding transition. Four isotopologues were synthesized to probe hydrogen bonding and solvent exposure of the beta-turn (K8), the N-terminus (S1), and the midstrand region (T10 and T3T10). Isotope-shifts, 2D lineshapes, and other spectral changes to the amide I 2D IR spectra of labeled TZ2 isotopologues were observed as a function of temperature. Data were interpreted on the basis of structure-based spectroscopic modeling of conformers obtained from extensive molecular dynamics simulations. The K8 spectra reveal two unique turn geometries, the type I' beta-turn observed in the NMR structure, and a less populated disordered or bulged loop. The data indicate that structures at low temperature resemble the folded NMR structure with typical cross-strand hydrogen bonds, although with a subpopulation of misformed turns. As the temperature is raised from 25 to 85 degrees C, the fraction of population with a type I' turn increases, but the termini also fray. Hydrogen bonding contacts in the midstrand region remain at all temperatures although with increasing thermal disorder. Our data show no evidence of an extended chain or random coil state for the TZ2 peptide at any temperature. The methods demonstrated here offer an approach to characterizing conformational variation within the folded or unfolded states of proteins and peptides.

Effect of viscogens on the kinetic response of a photoperturbed allosteric protein

“…2. That experimental observable evolves on roughly the same timescale as the C α (21)-C α (76) distance in the MD simulation and in a similar non-exponential fashion (it would be desirable to link the MD simulations more closely to the IR observable, as recently done for comparable protein systems, [53][54][55][56] which would be an interesting avenue for a future investigation). Fig.…”

Electrostatic frequency shifts in amide I vibrational spectra: Direct parameterization against experiment

“…[1][2][3] Delocalized amide I states provide sensitivity to secondary structure, while local peptide conformations can be probed by studying individual sites along the protein backbone, usually via isotope labels. [4][5][6][7] Although the rich information content of amide I spectra has been appreciated for many years, the complexity and congestion of the absorption spectra and the difficulty of accurately predicting the spectral features of individual protein structures have in many ways limited the utility of amide I spectroscopy in answering detailed structural questions. More recently, the advent of two-dimensional infrared (2DIR) spectroscopy has begun to assist in overcoming amide I spectral congestion, [8][9][10] while on the theoretical front, spectroscopic "maps" linking protein structure to specific spectral features have greatly improved our ability to interpret experimental data.…”

Solvent and conformation dependence of amide I vibrations in peptides and proteins containing proline

“…The 2DIR technique distinguishes β-sheet structures by producing Z shape like spectra [18,19] when this particular structural motif is present. In the past decades a number of 2DIR studies have been reported of peptides and proteins in solution [16,[20][21][22][23][24][25][26][27][28][29][30][31][32][33] or confined in membranes [34][35][36][37][38], revealing structural details and conformational changes from femtosecond (fs) to nanosecond (ns) time scales, and the nature of dynamic environments. For the structure determination of peptides that are in gas phase or micro-solvated (surrounded by few solvent molecules) the mid-infrared spectroscopic technique has become a promising tool [39][40][41][42].…”